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Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells
PURPOSE: gammaD-Crystallin (CRYGD) is a major structural lens crystallin and its mutations result in congenital cataract formation. In this study, we attempted to correct the altered protein features of G165fsX8 CRYGD protein with small chemical molecules. METHODS: Recombinant FLAG-tagged mutants (R...
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Formato: | Texto |
Lenguaje: | English |
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Molecular Vision
2010
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2890554/ https://www.ncbi.nlm.nih.gov/pubmed/20577655 |
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author | Gong, Bo Zhang, Li-Yun Lam, Dennis Shun-Chiu Pang, Chi-Pui Yam, Gary Hin-Fai |
author_facet | Gong, Bo Zhang, Li-Yun Lam, Dennis Shun-Chiu Pang, Chi-Pui Yam, Gary Hin-Fai |
author_sort | Gong, Bo |
collection | PubMed |
description | PURPOSE: gammaD-Crystallin (CRYGD) is a major structural lens crystallin and its mutations result in congenital cataract formation. In this study, we attempted to correct the altered protein features of G165fsX8 CRYGD protein with small chemical molecules. METHODS: Recombinant FLAG-tagged mutants (R15C, R15S, P24T, R61C, and G165fsX8) of CRYGD were expressed in COS-7 cells and treated with small chemical molecules with reported protein chaperoning properties (sodium 4-phenylbutyrate [4-PBA], trimethylamine N-oxide [TMAO], and glycerol and DMSO [DMSO]). Protein solubility in 0.5% Triton X-100 and subcellular distribution was examined by western blotting and immunofluorescence, respectively. Apoptosis was assayed as the percentage of fragmented nuclei in transfected cells. Expression of heat-shock proteins (Hsp70 and Hsp90) was examined by reverse transcription-polymerase chain reaction analysis. RESULTS: Unlike WT and most mutants (R15C, R15S, P24T, and R61C) of CRYGD, G165fsX8 CRYGD was significantly insoluble in 0.5% Triton X-100. This insolubility was alleviated by dose-dependent 4-PBA treatment. The treatment relieved the mislocalization of G165fsX8 CRYGD from the nuclear envelope. Also, 4-PBA treatment reduced cell apoptosis and caused an upregulation of Hsp70. CONCLUSIONS: 4-PBA treatment reduced the defective phenotype of mutant G165fsX8 CRYGD and rescued the affected cells from apoptosis. This could be a potential treatment for lens structural protein and prevent lens opacity in cataract formation. |
format | Text |
id | pubmed-2890554 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2010 |
publisher | Molecular Vision |
record_format | MEDLINE/PubMed |
spelling | pubmed-28905542010-06-24 Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells Gong, Bo Zhang, Li-Yun Lam, Dennis Shun-Chiu Pang, Chi-Pui Yam, Gary Hin-Fai Mol Vis Research Article PURPOSE: gammaD-Crystallin (CRYGD) is a major structural lens crystallin and its mutations result in congenital cataract formation. In this study, we attempted to correct the altered protein features of G165fsX8 CRYGD protein with small chemical molecules. METHODS: Recombinant FLAG-tagged mutants (R15C, R15S, P24T, R61C, and G165fsX8) of CRYGD were expressed in COS-7 cells and treated with small chemical molecules with reported protein chaperoning properties (sodium 4-phenylbutyrate [4-PBA], trimethylamine N-oxide [TMAO], and glycerol and DMSO [DMSO]). Protein solubility in 0.5% Triton X-100 and subcellular distribution was examined by western blotting and immunofluorescence, respectively. Apoptosis was assayed as the percentage of fragmented nuclei in transfected cells. Expression of heat-shock proteins (Hsp70 and Hsp90) was examined by reverse transcription-polymerase chain reaction analysis. RESULTS: Unlike WT and most mutants (R15C, R15S, P24T, and R61C) of CRYGD, G165fsX8 CRYGD was significantly insoluble in 0.5% Triton X-100. This insolubility was alleviated by dose-dependent 4-PBA treatment. The treatment relieved the mislocalization of G165fsX8 CRYGD from the nuclear envelope. Also, 4-PBA treatment reduced cell apoptosis and caused an upregulation of Hsp70. CONCLUSIONS: 4-PBA treatment reduced the defective phenotype of mutant G165fsX8 CRYGD and rescued the affected cells from apoptosis. This could be a potential treatment for lens structural protein and prevent lens opacity in cataract formation. Molecular Vision 2010-06-04 /pmc/articles/PMC2890554/ /pubmed/20577655 Text en Copyright © 2010 Molecular Vision. http://creativecommons.org/licenses/by/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Gong, Bo Zhang, Li-Yun Lam, Dennis Shun-Chiu Pang, Chi-Pui Yam, Gary Hin-Fai Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title | Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title_full | Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title_fullStr | Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title_full_unstemmed | Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title_short | Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells |
title_sort | sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammad-crystallin in cultured cells |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2890554/ https://www.ncbi.nlm.nih.gov/pubmed/20577655 |
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